title: Irinotecan synergistically enhances the antiproliferative and proapoptotic effects of axitinib in vitro and improves its anticancer activity in vivo.
abstract: OBJECTIVE
  To demonstrate the synergistic antiproliferative and proapoptotic activity of irinotecan and axitinib in vitro and the improvement of the in vivo effects on angiogenesis and pancreatic cancer.
  
  
  METHODS
  Proliferation and apoptotic assays were performed on human dermal microvascular endothelial cells and pancreas cancer (MIAPaCa-2, Capan-1) cell lines exposed to SN-38, the active metabolite of irinotecan, axitinib, or their simultaneous combination for 72 hours. ERK1/2 and Akt phosphorylation, the vascular endothelial growth factor (VEGF), VEGF receptor-2, and thrombospondin-1 (TSP-1) concentration were measured by ELISAs. ATP7A and ABCG2 gene expression was performed with real-time polymerase chain reaction and SN-38 intracellular concentrations were measured by high-performance liquid chromatography. Capan-1 xenografts in nude mice were treated with irinotecan and axitinib alone or in simultaneous combination.
  
  
  RESULTS
  A strong synergistic effect on antiproliferative and proapoptotic activity was found with the axitinib/SN-38 combination on endothelial and cancer cells. ERK1/2 and Akt phosphorylation were significantly inhibited by lower concentrations of the combined drugs in all the cell lines. Axitinib and SN-38 combined treatment greatly inhibited the expression of the ATP7A and ABCG2 genes in endothelial and cancer cells, increasing the SN-38 intracellular concentration. Moreover, TSP-1 secretion was increased in cells treated with both drugs, whereas VEGFR-2 levels significantly decreased. In vivo administration of the simultaneous combination determined an almost complete regression of tumors and tumor neovascularization.
  
  
  CONCLUSIONS
  In vitro results show the highly synergistic properties of simultaneous combination of irinotecan and axitinib on endothelial and pancreas cancer cells, suggesting a possible translation of this schedule into the clinics.
authors.affiliation: Division of Pharmacology, Department of Internal Medicine, University of Pisa, Pisa, Italy.
authors.forename: Bastianina
authors.initials: B
authors.lastname: Canu
authors.forename: Anna
authors.initials: A
authors.lastname: Fioravanti
authors.forename: Paola
authors.initials: P
authors.lastname: Orlandi
authors.forename: Teresa
authors.initials: T
authors.lastname: Di Desidero
authors.forename: Greta
authors.initials: G
authors.lastname: Alì
authors.forename: Gabriella
authors.initials: G
authors.lastname: Fontanini
authors.forename: Antonello
authors.initials: A
authors.lastname: Di Paolo
authors.forename: Mario
authors.initials: M
authors.lastname: Del Tacca
authors.forename: Romano
authors.initials: R
authors.lastname: Danesi
authors.forename: Guido
authors.initials: G
authors.lastname: Bocci
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects*; D000077784:Axitinib; D015153:Blotting, Western; D002166:Camptothecin / Q000008:administration & dosage / Q000031:analogs & derivatives; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D000073840:Copper-Transporting ATPases; D004357:Drug Synergism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007093:Imidazoles / Q000008:administration & dosage; D007124:Immunoenzyme Techniques; D007191:Indazoles / Q000008:administration & dosage; D000077146:Irinotecan; D008297:Male; D051379:Mice; D008819:Mice, Nude; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D009389:Neovascularization, Pathologic; D010190:Pancreatic Neoplasms / Q000098:blood supply / Q000473:pathology* / Q000517:prevention & control*; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019699:Thrombospondins / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
chemical_list: C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D027682:Cation Transport Proteins; D007093:Imidazoles; D007191:Indazoles; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D019699:Thrombospondins; D042461:Vascular Endothelial Growth Factor A; D000077146:Irinotecan; D000077784:Axitinib; D040301:Vascular Endothelial Growth Factor Receptor-2; D051057:Proto-Oncogene Proteins c-akt; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D000251:Adenosine Triphosphatases; C081184:ATP7A protein, human; D000073840:Copper-Transporting ATPases; D002166:Camptothecin